Francisella tularensis has been classified as a Category A biodefense agent by the Centers for Disease Control and Prevention because this bacterium is among the most virulent pathogens on the planet and causes a lethal infection if inhaled. Many pivotal studies in the field have demonstrated that replication within macrophages during infection is an important feature of the pathogenesis of F. tularensis. My laboratory has furthered the field by showing that interactions with non-macrophages are also extremely important during infection as these cells provide a niche for immune protection, proliferation, sanctuary from antibiotics, disease transmission, and other unexplored roles. Although much of the work in the field of F. tularensis has focused on the role of macrophages during infection, pathogen interactions with other cell types have not been thoroughly investigated. We have shown that F. tularensis invades and persists in erythrocytes. This invasion enhances resistance to antibiotics and enhances the ability of this bacterium to colonize ticks, important disease vectors. Preliminary data suggest that either the erythrocyte surface protein, Band 3, or the cytoskeletal protein, spectrin, are required for invasion. Further data show that a subset of bacterial genes induced in the presence of erythrocytes are important for invasion of these host cells. Moreover, structural proteins of the the bacterial type VI secretion system (T6SS) are required for invasion, suggesting that this apparatus secretes effector molecules into erythroctyes to mediate invasion. In this application, we will investigate bacterial genes induced in the presence of erythrocytes to provide further insight into the mechanism of red blood cell invasion. We will also identify the T6SS effector molecules that mediate this proces. Finally, we will determine whether Band 3, spectrin, or both host molecules are required for red blood cell invasion by F. tularensis.